Pancreatic cancer remains the fourth leading cause of cancer-related death in the United States with a median survival of less than 6 months following diagnosis. It is one of the most aggressive human malignancies with extremely poor prognosis, thus offering an experimental and clinical challenge. Rationale for a detailed pre-clinical evaluation of capsaicin for its efficacy against pancreatic cancer stems from our solid preliminary data from in vitro and xenograft studies. We showed that human pancreatic cancer cell lines AsPC-1 and BxPC-3 are highly sensitive to growth inhibition by capsaicin. Interestingly, the viability of acinar cells derived from normal human pancreas or immortalized normal human pancreatic ductal epithelial (HPDE-6) cells were minimally affected by capsaicin. These results are particularly encouraging because selective killing of cancer cells is a desirable characteristic of a potential cancer preventive or therapeutic agent. Our results also showed that growth inhibitory effect of capsaicin is associated with the (i) elevated levels of intracellular reactive oxygen species (ROS), (ii) increased expression of bax, (iii) leading to mitochondrial depolarization, release of cytochrome c and AIF, down regulation of survivin, and activation of caspase-3 cascade that eventually induces apoptosis in both AsPC-1 and BxPC-3 cells but not in acinar cells. In addition, we found that capsaicin significantly reduces glutathione and thioredoxin levels and activated JNK in pancreatic cancer cells, reflecting the potent alterations in intracellular redox triggered by the chili compound. Furthermore, we found that orally feeding capsaicin significantly retard the growth of AsPC-1 pancreatic tumor xenograft in nude mice with no discernible toxicity. Tumors obtained from capsaicin treated mice exhibited increased Bax expression, cytoplasmic cytochrome c, cleavage of caspase-3, reduced mitototic activity and increased apoptosis. Our results also showed that capsaicin enhances the growth suppressive effects of gemcitabine in pancreatic cancer cells. Building on these promising data, we hypothesize that capsaicin can effectively retard pancreatic carcinogenesis, and also exert potent anticancer effects against this tumor type, through a differential induction and modulation of oxidative stress leading to JNK activation and mitochondrial death pathway of apoptosis. To test this hypothesis we will: (1) Elucidate the mechanisms by which capsaicin triggers oxidative stress in pancreatic cancer cells, (2) Investigate the activation of mitogen- and stress-regulated signal transduction pathways in response to the oxidative imbalance induced by capsaicin, (3) Determine the effect of capsaicin on the activation of mitochondrial death pathway, and (4) Investigate the preventive and therapeutic effect of capsaicin in athymic nude mice and KrasG12D transgenic mice models of pancreatic intraepithelial neoplasia (PanINs) respectively and its bioavailability and pharmacokinetics. Successful completion of this project will support the development and clinical application of capsaicin for the chemoprevention of pancreatic cancer. PUBLIC HEALTH RELEVANCE: This translational research is to perform preclinical studies in pancreatic cancer cells using capsaicin, an active ingredient of chili pepper. We have provided lot of preliminary data to support that capsaicin is selective to cancer cells and is not toxic to normal cells. We propose to determine its molecular mechanism in culture and in the animal model.